Epithelial Necrosis Research Articles

A novel herpes-like virus inducing branchial lesions in a tiger shark (Galeocerdo cuvier).

A juvenile, male tiger shark (Galeocerdo cuvier) developed illness after capture in Florida waters and was euthanized. Gross lesions included mild skin abrasions, hepatic atrophy, and coelomic fluid. Histologically, gills contained multifocal lamellar epithelial cell necrosis and thromboses. Scattered gill and esophageal epithelial cells had large, basophilic, intracytoplasmic, and intranuclear inclusions. Ultrastructurally, lamellar epithelial cells contained arrays of intracytoplasmic viral particles and scattered intranuclear nucleocapsids. Capsulated virions were 148 ± 11 nm with an 84 ± 8 nm icosahedral nucleocapsid and an electron-dense core. Next-generation sequencing, quantitative polymerase chain reaction, and in situ hybridization performed on formalin-fixed tissue confirmed a herpes-like viral infection. The viral polymerase shared 24% to 31% protein homology with other alloherpesviruses of fish, indicating a divergent virus. This report documents the pathologic findings associated with a molecularly confirmed novel herpes-like virus in an elasmobranch.

Pirfenidone Attenuates Renal Tubulointerstitial Fibrosis through Inhibiting miR-21

Background: Our previous studies had shown pirfenidone (PFD) not only improved tubulointerstitial fibrosis (TIF) but also inhibited the expression of microRNA-21 (miR-21) in the renal tissue of unilateral urethral obstruction (UUO) rats. This study aims to investigate whether PFD can attenuate TIF through inhibiting miR-21 in UUO rats. Methods: Sprague Dawley rats were divided randomly into sham-operated group, UUO group, and PFD and olmesartan (Olm) treatment groups. Samples were collected on day 14. Expression of miR-21, TGF-β1, Smad3, and Smad7 mRNA in the renal tissue was detected using real-time quantitative PCR. Immunohistochemistry was performed to assess the protein expressions of collagen III, E-cadherin, and α-SMA. Automated capillary Western blotting was used to detect the quantitative expression of TGF-β1, Smad3, p-Smad3, Smad7, collagen III, E-cadherin, and α-SMA in renal tissues. The expression of miR-21 and Smad7 mRNA and the protein levels of collagen III and α-SMA were examined in the miR-21-overexpressing cell line, NRK-52E. Results: Compared with the UUO group, both PFD and Olm inhibited renal tubular dilation, diffused epithelial cell degeneration and necrosis, and reduced renal interstitial edema, inflammatory cell infiltration, and collagen fiber deposition, while no significant difference between PFD group and Olm group. Informatics-based approaches identified Smad7 as a likely candidate for regulation by miR-21. Compared with the sham group, miR-21 expression was upregulated in the UUO group resulting in the downregulation of Smad7 expression due to degradation. The overexpression of miR-21 in the in vitro model downregulated Smad7 and promoted EMT and ECM accumulation. Protein levels of TGF-β1, Smad3, p-Smad3, collagen III, and α-SMA were upregulated, while E-cadherin protein was downregulated in the UUO group than in the sham group. PFD rather than Olm decreased the expression of miR-21 and increased the expression level of Smad7 mRNA and then inhibited the TGF-β1/Smad3 signaling pathway. Olm only downregulated the TGF-β1/Smad3 signaling pathway. Conclusions: PFD improves TIF by downregulating the expression of miR-21, then elevating Smad7, and finally inhibiting the activation of the TGF-β1/Smad3 signaling pathway in UUO rats.

Histopathological and immunohistochemical study of periodontal changes in chronic smokers.

Periodontal disease is a chronic inflammatory, multifactorial condition, that, in the absence of an early and adequate treatment, may lead to a progressive damaging of the alveolar tissues that support the teeth (periodontal ligament, cement and alveolar bone) followed by teeth mobility and, subsequently, their loss. Periodontal disease is one of the most common inflammatory disease affecting adult individuals all over the world, being considered a real worldwide pandemic. This disease may influence the progression of certain systemic diseases: diabetes mellitus, cardiovascular diseases, ischemic cardiomyopathy, myocardial infarction, stroke, neurodegenerative diseases, chronic kidney diseases, cancer, etc. The association between smoking and periodontal disease was described in numerous clinical and epidemiological studies, suggesting that products derived from tobacco burning may change the clinical aspects and the disease progression. The present study analyzed microscopically and immunohistochemically 58 periodontal fragments, from 50 patients, chronic smokers, clinically diagnosed with severe periodontitis. There were highlighted major changes in the gingival epithelium (epithelium thickening, acanthosis, intraepithelial edema, infiltrates of neutrophils or lymphocytes, epithelial necrosis), in the periodontal conjunctive tissue (more or less intense inflammatory infiltrates, microhemorrhages, vascular congestion, intense immunohistochemical expression for some matrix metalloproteinases). The periodontal changes may be the expression of both toxic factors present in tobacco smoke and due to the changes caused by tobacco in the microbial flora of the oral cavity.

Enterogyrus spp. (Monogenea: Ancyrocephalinae) and Aeromonas jandaei co-infection associated with high mortality following transport stress in cultured Nile tilapia.

Monogenean infection of the internal organs is extremely rare when compared to external infections. This study describes mass mortality of Nile tilapia (Oreochromis niloticus L.) originating from co-infection with Enterogyrus spp. and Aeromonas jandaei following transport stress. The first fish deaths occurred on day 1 post-transport, while cumulative mortality reached approximately 90% by day 10 post-stocking. An atypical amount of pale (whitish) faeces floating on the surface of the water as well as typical clinical signs of motile Aeromonas septicemia, were reported. Adult monogeneans and countless eggs of monogeneans were found in the stomachs and the intestines of both moribund and dead fish, respectively. Two strains of A. jandaei were isolated from the kidneys. Scanning electron microscope microphotographs of the stomach revealed the presence of numerous monogeneans penetrating deep into the gastric tissue, and diffuse lesions filled with bacilliform bacteria. Histopathological examination showed multifocal eosinophilic infiltrate, gastric gland and epithelial necrosis with sloughed necrotic debris in the lumen. This is the first report of co-infection by Enterogyrus spp. and A. jandaei in Nile tilapia and the first report of Enterogyrus coronatus, Enterogyrus foratus, and Enterogyrus malbergi parasitizing tilapia in Brazil. These findings indicate that synergic co-infection by Monogenean stomach parasites (E. coronatus, E. foratus, and E. malbergi) and A. jandaei may induce high mortalities in tilapia following transport stress.

Hydrogen-Rich Water Alleviates the Nickel-Induced Toxic Responses (Inflammatory Responses, Oxidative Stress, DNA Damage) and Ameliorates Cocoon Production in Earthworm.

In recent years, studies investigating the protective effect of hydrogen-rich water (HRW) against different diseases and the toxicity of some substances have attracted increasing attention. Here, we assessed the effects of hydrogen-rich water on different nickel-induced toxic responses (reactive oxygen species (ROS), tumor necrosis factor-alpha (TNF-α), and 8-hydroxy-2'-deoxyguanosine (8-OHdG) of stress responses, histopathological changes) and cocoon production in earthworm model. Earthworms were randomly divided into two main groups: water (W) group including control (CW: ultrapure water), 10 (10W), 200 (200W), and 500 (500W), and hydrogen-rich ultrapure water (HRW) group including control (CHRW: hydrogen-rich ultrapure water), 10 (10HRW), 200 (200HRW), and 500 (500HRW) mg of nickel chloride kg-1 soil for 14days. We found that cocoon production was less affected by the nickel exposure of earthworms in the 500HRW group compared to the 500W group. The ROS levels in 200HRW and 500HRW groups were less than that of 200W and 500W, respectively. The epithelial degeneration, epithelial necrosis, and necrosis in muscle fibers in tissues of earthworm were less damaged in 200HRW and 500HRW groups compared to 200W and 500W, respectively. HRW groups significantly reduced the expression of 8-OHdG induced by nickel exposure and inflammatory cytokine response including TNF-α. The study showed that hydrogen-rich water could alleviate the toxic effects of nickel-induced oxidative and inflammatory damages in earthworms. The HRW treatment known for its cheap and eco-friendly propertıes without any negative effects on the ecosystem can be used as a green method for alleviating the toxification effects of heavy metals in contaminated soil and increasing cocoon production of earthworms.

Multi-Organ Histopathological Changes in a Mouse Hepatitis Virus Model of COVID-19

Infection with SARS-CoV-2, the virus responsible for the global COVID-19 pandemic, causes a respiratory illness that can severely impact other organ systems and is possibly precipitated by cytokine storm, septic shock, thrombosis, and oxidative stress. SARS-CoV-2 infected individuals may be asymptomatic or may experience mild, moderate, or severe symptoms with or without pneumonia. The mechanisms by which SARS-CoV-2 infects humans are largely unknown. Mouse hepatitis virus 1 (MHV-1)-induced infection was used as a highly relevant surrogate animal model for this study. We further characterized this animal model and compared it with SARS-CoV-2 infection in humans. MHV-1 inoculated mice displayed death as well as weight loss, as reported earlier. We showed that MHV-1-infected mice at days 7–8 exhibit severe lung inflammation, peribronchiolar interstitial infiltration, bronchiolar epithelial cell necrosis and intra-alveolar necrotic debris, alveolar exudation (surrounding alveolar walls have capillaries that are dilated and filled with red blood cells), mononuclear cell infiltration, hyaline membrane formation, the presence of hemosiderin-laden macrophages, and interstitial edema. When compared to uninfected mice, the infected mice showed severe liver vascular congestion, luminal thrombosis of portal and sinusoidal vessels, hepatocyte degeneration, cell necrosis, and hemorrhagic changes. Proximal and distal tubular necrosis, hemorrhage in interstitial tissue, and the vacuolation of renal tubules were observed. The heart showed severe interstitial edema, vascular congestion, and dilation, as well as red blood cell extravasation into the interstitium. Upon examination of the MHV-1 infected mice brain, we observed congested blood vessels, perivascular cavitation, cortical pericellular halos, vacuolation of neuropils, darkly stained nuclei, pyknotic nuclei, and associated vacuolation of the neuropil in the cortex, as well as acute eosinophilic necrosis and necrotic neurons with fragmented nuclei and vacuolation in the hippocampus. Our findings suggest that the widespread thrombotic events observed in the surrogate animal model for SARS-CoV-2 mimic the reported findings in SARS-CoV-2 infected humans, representing a highly relevant and safe animal model for the study of the pathophysiologic mechanisms of SARS-CoV-2 for potential therapeutic interventions.

Mortality of Clarias gariepinus caused by Aeromonas caviae and nitrite toxicity in a fish farm

In Nigeria, farming of freshwater fish, mainly that of Clarias gariepinus has gained prominence as means of improved protein supply and livelihood. Many farmers suffer untold losses in their bid to make a living from commercial fish production. Among the major causes of infectious disease outbreaks in fish farms are pathogenic bacteria of the genus Aeromonas. This is a case report of outbreak of Aeromonas caviae infection complicated by nitrite toxicity in Clarias gariepinus. Carcasses of 17 African catfish juveniles and two water samples were presented to the poultry and fish clinic of the Veterinary Teaching Hospital, University of Jos for investigation. The fishes had been on medication (Fish cure antibiotic +®) for over 5 days with cumulative mortality of 230 out of 2,500 fishes (9.2 %). Necropsy was conducted and samples were taken for microbial analysis and histopathology while water samples were subjected to chemical analysis. Gross lesions were broken barbels, cutaneous depigmentation, branchial pallor, ascites, renal and splenic congestion. Histologically, there was dissociation of hepatic cords, necrosis of hepatocytes with vacuolation and mononuclear cellular infiltrations. Also, there was renal tubular epithelial necrosis with mononuclear and heterophilic infiltration while micro abscesses were observed in the brain. Marked Zenker’s necrosis and edema were seen in the skeletal muscles. Organism isolated from livers and kidneys was identified as Aeromonas caviae and was susceptible to enrofloxacin, furaltadone and florfenicol. Nitrite in fish pond water was 1mg/L. Bath medication with enrofloxacin at 33mg/litre of water for 6 hours treatment per day was done to reduce the mortality. The farmer was advised to change the source of water for the fish pond immediately. It is recommended that to avoid losses in fish farming, farmers should check the quality of water intended for use on fish farms and ensure early health check on fingerlings purchased for rearing.

Effect of virgin low density polyethylene microplastic ingestion on intestinal histopathology and microbiota of gilthead sea bream

Polyethylene microparticles are common contaminants in the marine environment, which are easily ingested by fish and can provoke physical damage or toxic reactions, resulting in inflammation, stress-related alterations and behavioural disorders. This work focuses on the extend of the alterations by microplastic ingestion on the intestinal health of Sparus aurata, in an integrative effort to relate the effect on gut microbiota and histopathological damage. Virgin, low density, polyethylene microplastic (LDPE-MP) particles were included into a control diet (D1, without MP), at a 0.12% in two different size ranges in diet D2 (200–500 μm) and D3 (501–1000 μm). Gilthead sea bream were fed D1, D2 or D3 diets for 30 days, and then sacrificed, biometrical data recorded, anterior intestine samples taken for histological analyses and anterior intestine mucus collected for Illumina sequencing of the V3-V4 region of the 16S rRNA, to evaluate the adherent bacterial communities. Intestines of fish fed D2 and D3 contained plastic aggregates and showed macroscopic signs of distension, liquid accumulation and inflammation. Their histopathological analysis revealed severe mechanical lesions (haemorrhages and epithelial desquamation and necrosis), inflammatory reaction (eosinophilia, lymphocytosis, and melanomacrophage presence) and severe epithelial vacuolization, caused by erosion and accumulation of MPs with sharp and rough-edges. Illumina sequencing resulted in over 7 million high-quality reads that were assigned to 589 OTUs. No significant differences were found in richness, diversity or composition at the phylum level among all three experimental groups. However, small but significant differences were found in differential expression analyses, with the D2 diet changing 6.2% and D3 changing 2.9% of the total bacterial composition in anterior intestine. The altered microbial composition seems to be directed as an attempt to counteract the mechanical damage and inflammation inflicted by MPs ingestion, in agreement with metagenome prediction and pathway analysis, which revealed that the slight effect of diets containing MPs (D2 and D3) on gut microbiota has the potential to change the metabolic capacity of these populations, including a decrease in pathways related to bacterial infection and inflammation. Thus, virgin LDPE-MPs ingestion in gilthead sea bream did not significantly affect growth and gut microbiota composition over a 30-day feeding period, but the slight alterations observed could derive in growth problems and dysbiosis upon prolonged microplastic ingestion, which are mainly driven by the severe mechanical damage inflicted to the digestive tract.

A High Starch Diet Alters the Composition of the Intestinal Microbiota of Largemouth Bass Micropterus salmoides, Which May Be Associated With the Development of Enteritis.

Starch is an inexpensive feed ingredient that has been widely used in fish feed. However, starch utilization by carnivorous fish is limited and excess starch is detrimental to the health of the organism. High starch diets often lead to liver damage, but the effects on the intestine are often overlooked. Therefore, in this study, two isonitrogenous and isolipidic semi-pure diets (NC: 0% α-starch, HC: 22% α-starch) were formulated and fed to largemouth bass (Micropterus salmoides) for 45 days. The effects of the high starch diet on the intestine of largemouth bass were comprehensively investigated by intestinal microbiota, histopathology, ultrastructural pathology, and enzymology analyses. Feeding the HC diet did not affect the growth of largemouth bass during the experimental period. However, the high starch diet led to a reduction in the diversity and abundance of intestinal microbiota in largemouth bass, with a significant increase in the abundance of harmful bacteria (Aeromonas) and a decrease in the abundance of beneficial bacteria (Clostridium, Lactobacillus, and Bifidobacterium). Feeding the HC diet caused the development of enteritis, with goblet cell hyperplasia, epithelial necrosis and detachment and inflammatory cell infiltration, and leading to enlarged apical openings and mitochondrial damage in goblet cells. Long-term feeding of the HC diet inhibited intestinal α-amylase activity. changes in the intestinal microbiota, such as an increase in Aeromonas and a decrease in Clostridium, Lactobacillus, and Bifidobacterium, may be closely related to the development of enteritis. Therefore, adding these beneficial bacteria as probiotics may be an effective way to prevent damage to the intestine of largemouth bass from a high carbohydrate diet. Our results suggest reducing the amount of starch added to the largemouth bass diets. This study provides a reference for protecting the largemouth bass gut during modern intensive culture.

The 1918 influenza and COVID-19 pandemics: The effect of age on outcomes.

The 1918 influenza and COVID-19 pandemics: The effect of age on outcomes.

Candidalysin triggers epithelial cellular stresses that induce necrotic death

Candida albicans is a common opportunistic fungal pathogen that causes a wide range of infections from superficial mucosal to hematogenously disseminated candidiasis. The hyphal form plays an important role in the pathogenic process by invading epithelial cells and causing tissue damage. Notably, the secretion of the hyphal toxin candidalysin is essential for both epithelial cell damage and activation of mucosal immune responses. However, the mechanism of candidalysin-induced cell death remains unclear. Here, we examined the induction of cell death by candidalysin in oral epithelial cells. Fluorescent imaging using healthy/apoptotic/necrotic cell markers revealed that candidalysin causes a rapid and marked increase in the population of necrotic rather than apoptotic cells in a concentration dependent manner. Activation of a necrosis-like pathway was confirmed since C. albicans and candidalysin failed to activate caspase-8 and -3, or the cleavage of poly (ADP-ribose) polymerase. Furthermore, oral epithelial cells treated with candidalysin showed rapid production of reactive oxygen species, disruption of mitochondria activity and mitochondrial membrane potential, ATP depletion and cytochrome c release. Collectively, these data demonstrate that oral epithelial cells respond to the secreted fungal toxin candidalysin by triggering numerous cellular stress responses that induce necrotic death. TAKE AWAYS: Candidalysin secreted from Candida albicans causes epithelial cell stress. Candidalysin induces calcium influx and oxidative stress in host cells. Candidalysin induces mitochondrial dysfunction, ATP depletion and epithelial necrosis. The toxicity of candidalysin is mediated from the epithelial cell surface.

Il4ra-independent vaginal eosinophil accumulation following helminth infection exacerbates epithelial ulcerative pathology of HSV-2 infection

SummaryHow helminths influence the pathogenesis of sexually transmitted viral infections is not comprehensively understood. Here, we show that an acute helminth infection (Nippostrongylus brasiliensis [Nb]) induced a type 2 immune profile in the female genital tract (FGT). This leads to heightened epithelial ulceration and pathology in subsequent herpes simplex virus (HSV)-2 infection. This was IL-5-dependent but IL-4 receptor alpha (Il4ra) independent, associated with increased FGT eosinophils, raised vaginal IL-33, and enhanced epithelial necrosis. Vaginal eosinophil accumulation was promoted by IL-33 induction following targeted vaginal epithelium damage from a papain challenge. Inhibition of IL-33 protected against Nb-exacerbated HSV-2 pathology. Eosinophil depletion reduced IL-33 release and HSV-2 ulceration in Nb-infected mice. These findings demonstrate that Nb-initiated FGT eosinophil recruitment promotes an eosinophil, IL-33, and IL-5 inflammatory circuit that enhances vaginal epithelial necrosis and pathology following HSV-2 infection. These findings identify a mechanistic framework as to how helminth infections can exacerbate viral-induced vaginal pathology.

Coadministration of Ginger Extract and Fluconazole Shows a Synergistic Effect in the Treatment of Drug-Resistant Vulvovaginal Candidiasis.

BackgroundAzoles are the most common antifungal drugs used in the treatment of vulvovaginal candidiasis (VVC). The frequency of azole-resistant Candida isolates has increased dramatically in the last two decades. Here, we assessed the antifungal activity of a combination of fluconazole (FLZ) and methanolic extract of ginger (Meth-Gin) against drug-resistant vulvovaginal candidiasis (VVC) in a murine model.MethodsThe in vitro activity of FLZ or a combination of FLZ and Meth-Gin was determined against Candida albicans by the agar well diffusion, macrodilution, time-kill and the biofilm eradication methods. The therapeutic efficacy of the formulations was assessed by analyzing the fungal load, pro-inflammatory cytokines, percent apoptotic cells and the histological changes in the vaginal tissues of the mice. Moreover, the renal toxicity the drug formulation was evaluated by analyzing the levels of the blood urea nitrogen (BUN) and creatinine.ResultsThe results of in vitro study demonstrated that FLZ did not show any activity against C. albicans, whereas a combination of FLZ and Meth-Gin demonstrated greater activity as shown by the data of the zone of growth inhibition, MIC and time-kill assay. FLZ or Meth-Gin treatment could not completely cure VVC, whereas a combination of FLZ and Meth-Gin was greatly effective in the treatment of VVC. The vaginal tissue from mice of the infected control group had the highest fungal load of 155370 ± 20617 CFUs. Treatment with FLZ at a dose of 40 mg/kg reduced the fungal load to 120863 ± 10723 CFUs. Interestingly, the mice treated with a combination of FLZ (40 mg/kg) and Meth-Gin (200 mg/kg) had a fungal load of 256 ± 152 CFUs. Besides, FLZ and Meth-Gin combination effectively reduced the pro-inflammatory cytokines (IL-1β, TNF-α and IL-17) and the percentage of apoptotic cells in the vaginal tissues. Likewise, the histological analysis revealed the epithelial necrosis, shedding and ulceration in the vaginal tissue, whereas treatment with FLZ and Meth-Gin combination reversed the histopathological changes in the vaginal epithelium and lamina propria.ConclusionThe findings of the current study suggest that the co-administration of Meth-Gin and FLZ may have a potential therapeutic effect in the treatment of azole-resistant candidiasis.

Retrospective and prospective studies of transmissible viral proventriculitis in broiler chickens in Brazil

We investigated the occurrence and pathologic findings of transmissible viral proventriculitis (TVP) associated with the chicken proventricular necrosis virus (CPNV) in commercial broiler chickens in southeastern Brazil. Seventy-three broilers, 25-36 d old, with a history of reduced growth, were referred to our veterinary pathology services from 2013 to 2017. Broilers were clinically examined, weighed, and euthanized for postmortem examination. Broilers of different ages with proventricular histologic lesions were positive for CPNV by RT-PCR; however, the intensity of histologic lesions was higher among 33-d-old animals, and viral RNA detection was more frequent among those that were 28 d old. In the proventriculi of 35 of 73 (48%) broilers, lesions were characterized by glandular epithelial necrosis, lymphoplasmacytic and histiocytic infiltrates, and metaplasia of glandular epithelium to ductal epithelium. In 24 of 73 (36%) broilers with histologic TVP-compatible lesions, CPNV was detected by RT-PCR for the viral protein 1 (VP1) gene. Broilers with histologic lesions were lighter than expected compared to the Cobb 500 standard weight. TVP has not been reported previously in broiler chickens in Brazil, to our knowledge.

Study on baicalin on liver and kidney toxicity in male rats

Objective: To observe the toxic effects of different doses of baicalin on liver and kidney of rats after different time administration, and provide experimental reference for the safety of clinical medication. Methods: In April 2019, 42 Wistar male rats were randomly divided into a control group (0.9% sodium chloride solution) and baicalin administration groups (100, 200 mg/kg) , 14 rats in each group, and one was given by oral gavage. 7 times/d, 6 times/week, 7 rats in each group were sacrificed 28 and 56 days after the administration. The wet weights of liver and kidney were weighed and the organ coefficients were calculated. The hematoxylin-eosin (HE) staining was used to detect the histomorphological changes. And the levels of serum alanine aminotransferase (ALT) , aspartate aminotransferase (AST) , alkaline phosphatase (ALP) , blood urea nitrogen (BUN) and creatinine (CRE) were detected. Results: After 56 days of administration in baicalin 200 mg/kg rats, the body weight and kidney coefficient were lower than those of the control group. Histopathology showed that glomerular atrophy became smaller, renal tubules were significantly atrophied, and epithelial cell necrosis occurred. No obvious abnormalities in liver was observed. After 56 days of administration in baicalin 200 mg/kg rats, the levels of BUN and CRE in the serum were higher than those of the control group, and the differences were statistically significant (P<0.05) . There were no obvious abnormalities in the baicalin 100 mg/kg group and the 28 d of administration in baicalin 200 mg/kg group. Conclusion: Under the conditions of this test, baicalin has certain renal toxicity in male rats.

Viral tropism and detection of clade 2.3.4.4b H5N8 highly pathogenic avian influenza viruses in feathers of ducks and geese

Highly Pathogenic Avian Influenza viruses (HPAIVs) display a tissue pantropism, which implies a possible spread in feathers. HPAIV detection from feathers had been evaluated for H5N1 or H7N1 HPAIVs. It was suggested that viral RNA loads could be equivalent or higher in samples of immature feather compared to tracheal (TS) or cloacal swabs (CS). We investigated the suitability of feathers for the detection of clade 2.3.4.4b H5N8 HPAIV in ducks and geese field samples. In the six H5N8 positive flocks that were included in this study, TS, CS and immature wing feathers were taken from at least 10 birds. Molecular loads were then estimated using real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR) targetting H5 and M genes. In all flocks, viral loads were at least equivalent between feather and swab samples and in most cases up to 103 higher in feathers. Bayesian modelling confirmed that, in infected poultry, RT-qPCR was much more likely to be positive when applied on a feather sample only (estimated sensitivity between 0.89 and 0.96 depending on the positivity threshold) than on a combination of a tracheal and a cloacal swab (estimated sensitivity between 0.45 and 0.68 depending on the positivity threshold). Viral tropism and lesions in feathers were evaluated by histopathology and immunohistochemistry. Epithelial necrosis of immature feathers and follicles was observed concurrently with positive viral antigen detection and leukocytic infiltration of pulp. Accurate detection of clade 2.3.4.4b HPAIVs in feather samples were finally confirmed with experimental H5N8 infection on 10-week-old mule ducks, as viral loads at 3, 5 and 7 days post-infection were higher in feathers than in tracheal or cloacal swabs. However, feather samples were associated with lower viral loads than tracheal swabs at day 1, suggesting better detectability of the virus in feathers in the later course of infection. These results, based on both field cases and experimental infections, suggest that feather samples should be included in the toolbox of samples for detection of clade 2.3.4.4b HPAI viruses, at least in ducks and geese.

Safety and efficacy of combination of suberoylamilide hydroxyamic acid and mitomycin C in reducing pro-fibrotic changes in human corneal epithelial cells

Corneal haze post refractive surgery is prevented by mitomycin c (MMC) treatment though it can lead to corneal endothelial damage, persistent epithelial defects and necrosis of cells. Suberanilohydroxamic acid (SAHA) however has been proposed to prevent corneal haze without any adverse effects. For clinical application we have investigated the short and long term outcome of cells exposed to SAHA. Human donor cornea, cultured limbal epithelial cells, corneal rims and lenticules were incubated with SAHA and MMC. The cells/tissue was then analyzed by RT-qPCR, immunofluorescence and western blot for markers of apoptosis and fibrosis. The results reveal that short term exposure of SAHA and SAHA + MMC reduced apoptosis levels and increased αSMA expression compared to those treated with MMC. Epithelial cells derived from cultured corneal rim that were incubated with the MMC, SAHA or MMC + SAHA revealed enhanced apoptosis, reduced levels of CK3/CK12, ∆NP63 and COL4A compared to other treatments. In SAHA treated lenticules TGFβ induced fibrosis was reduced. The results imply that MMC treatment for corneal haze has both short term and long term adverse effects on cells and the cellular properties. However, a combinatorial treatment of SAHA + MMC prevents expression of corneal fibrotic markers without causing any adverse effect on cellular properties.

Prophylactic and Therapeutic Role of Vitamin D3 in Combination with Fluconazole Against Vaginal Candidiasis in a Murine Model.

In the present study, we assessed the adjunct effect of vitamin D3 in combination with Fluconazole (FLZ) against Vulvovaginal Candidiasis (VVC) in mice. Prophylactic effect was assessed by pretreating mice with vitamin D3 before exposure of mice with 2 X 106 CFUs of Candida albicans followed by treatment with FLZ. To determine the combined therapeutic efficacy, C. albicans infected mice were treated with a combination of vitamin D3 (10 μg/kg) and FLZ (10 and 20 mg/kg). The efficacy of the treatment was assessed by analyzing the fungal load and blood cell count. Moreover, the levels of inflammatory cytokines, including IL- 1β, IL-17 and TNF-α, were analyzed in the vaginal tissues. The histological analysis of the vaginal tissue from the untreated and treated mice was performed to assess the efficacy of the treatment. Prophylactic treatment with vitamin D3 (10 and 20 μg/kg) significantly increased the therapeutic effect of FLZ against VVC. In a therapeutic experiment, mice in the infected control group showed the highest vaginal fungal load of 83627 ± 10058 CFUs. Treatment with FLZ at a dose of 10 mg/kg reduced fungal load to 55523 ± 14823 CFUs, whereas the mice treated with a combination of vitamin D3 and FLZ (10 mg/kg) had the fungal burden of 12156 ± 3219. Similarly, treatment with FLZ (20 mg/kg) reduced fungal load to 36394 ± 5648 CFUs, whereas the addition of vitamin D3 to FLZ (20 mg/kg) further reduced the fungal burden to 2179 ± 1188. The leukocyte numbers in the infected mice increased to 9802 ± 505 as compared to 5152 ± 778 in normal control mice. Whereas, a combination of vitamin D3 with FLZ (10 and 20 mg/kg) reduced leukocyte numbers to 7284 ± 607 and 5739 ± 1126. The histological analysis data revealed epithelial necrosis, shedding and ulceration in the vaginal wall. Treatment with FLZ or a combination of FLZ and vitamin D3 brought regenerative changes in the vaginal epithelium and lamina proparia. The results of the present work recommend that the addition of vitamin D3 may be considered to increase the efficacy of FLZ against VVC.

Pulmonary and vascular effects of acute ozone exposure in diabetic rats fed an atherogenic diet

Air pollutants may increase risk for cardiopulmonary disease, particularly in susceptible populations with metabolic stressors such as diabetes and unhealthy diet. We investigated effects of inhaled ozone exposure and high-cholesterol diet (HCD) in healthy Wistar and Wistar-derived Goto-Kakizaki (GK) rats, a non-obese model of type 2 diabetes. Male rats (4-week old) were fed normal diet (ND) or HCD for 12 weeks and then exposed to filtered air or 1.0 ppm ozone (6 h/day) for 1 or 2 days. We examined pulmonary, vascular, hematology, and inflammatory responses after each exposure plus an 18-h recovery period. In both strains, ozone induced acute bronchiolar epithelial necrosis and inflammation on histopathology and pulmonary protein leakage and neutrophilia; the protein leakage was more rapid and persistent in GK compared to Wistar rats. Ozone also decreased lymphocytes after day 1 in both strains consuming ND (~50%), while HCD increased circulating leukocytes. Ozone increased plasma thrombin/antithrombin complexes and platelet disaggregation in Wistar rats on HCD and exacerbated diet effects on serum IFN-γ, IL-6, KC-GRO, IL-13, and TNF-α, which were higher with HCD (Wistar>GK). Ex vivo aortic contractility to phenylephrine was lower in GK versus Wistar rats at baseline(~30%); ozone enhanced this effect in Wistar rats on ND. GK rats on HCD had higher aortic e-NOS and tPA expression compared to Wistar rats. Ozone increased e-NOS in GK rats on ND (~3-fold) and Wistar rats on HCD (~2-fold). These findings demonstrate ways in which underlying diabetes and HCD may exacerbate pulmonary, systemic, and vascular effects of inhaled pollutants.

An Unrecognized Hazard in E-Cigarette Vapor: Preliminary Quantification of Methylglyoxal Formation from Propylene Glycol in E-Cigarettes.

Up to 95% of the liquid volume in an e-cigarette consists of propylene glycol. Previous research has shown that propylene glycol can generate diacetyl and formaldehyde when heated. New research shows that propylene glycol can also generate methylglyoxal, an alpha di-carbonyl compound recently shown to cause epithelial necrosis at even lower concentrations than diacetyl, the flavoring chemical associated with bronchiolitis obliterans (“Popcorn Lung”). We analyzed chemical emissions from 13 JUUL pod flavors. Diacetyl and methylglyoxal was detected in 100% of samples with median concentration (range) of 20 µg/m3 (less than limit of quantification: 54 µg/m3) and 4219 µg/m3 (677–15,342 µg/m3), respectively. We also detected acetaldehyde (median concentration: 341 µg/m3) and propionaldehyde (median concentration: 87 µg/m3) in all samples. The recent evidence that methylglyoxal is more cytotoxic to airway epithelial cells than diacetyl makes this an urgent public health concern. Current smokers considering e-cigarettes as a smoking cessation tool, and never users, who may be under the impression that e-cigarettes are harmless, need information on emissions and potential risks to make informed decisions.